Figure 3.

Bone-targeted AAV gene silencers reverse bone loss in osteoporosis

(A) mRNA levels of Shn3 or Sost in the tibia of young (2-month-old) or old (20-month-old) mice (n = 5). (B) Diagram of the study and treatment methods. (C–E) Sham or OVX surgery was performed on 3-month-old female mice, and, 6 weeks later, mice were i.v. injected with rAAV9.DSS (5 × 10¹³ vg/kg) carrying amiR-ctrl, amiR-shn3, amiR-sost, or amiR-sost/shn3. Eight weeks later, mRNA levels of Shn3 and Sost in the tibia were assessed by RT-PCR and normalized to Hprt (C, n = 9). Femoral bone mass was assessed by microCT (D and E). Representative 3D-reconstruction (D) and relative quantification (E) are displayed (n = 5–10). Scale bar: (D) 1 mm. (F–I) Twenty-month-old male mice were i.v. injected with rAAV9.DSS (5 × 10¹³ vg/kg) carrying amiR-ctrl, amiR-shn3, amiR-sost, or amiR-sost/shn3, and, 2 months later, mRNA levels of Shn3 and Sost in the tibia were assessed by RT-PCR and normalized to Hprt (F, n = 6). Trabecular bone mass in femurs (G and H) and lumbar vertebrae (L4, I) were assessed by microCT. Representative 3D reconstruction (G) and relative quantification (H and I) are displayed (n = 8–10). Scale bar: (G) 1 mm. (J and K) Sham or OVX surgery was performed on 3-month-old female mice, and, 6 weeks later, mice were i.v. injected with rAAV9.DSS. Histomorphometric quantification of BFR/BS and MAR was performed 8 weeks post injection (J). Serum CTX-I levels were measured to assess in vivo osteoclast activity (K). Values represent mean ± SD by an unpaired two-tailed Student’s t test (A) and one-way ANOVA test (C, E, F, H–K).